Project Summary The nerve terminals of auditory neurons are responsible for the efficient release and recycling of synaptic vesicles that enable the secure chemical transmission that underlies auditory perception. The fidelity and modulation of these synaptic events is important in the superior olivary complex (SOC) of the mammalian brainstem, which is involved in sound source localization. The calyx of Held nerve terminal is an integral component of this afferent projection pathway, which projects from the globular bushy cells (GBCs) of the anteroventral cochlear nucleus (aVCN) and synapses onto the principal cells (PCs) of the medial nucleus of the trapezoid bodies (MNTB). The calyx of Held-MNTB synapse contributes specifically to the transduction of interaural intensity differences from sound cues initiated at each cochlea. Although it is well-established that synaptic efficacy changes at an acute timescale in response to previous activity, the underlying mechanisms regulating short-term modifications in synaptic strength in the auditory brainstem require further investigation. The work proposed here seeks to understand how retrograde signaling regulates presynaptic short-term plasticity (STP) at the mouse calyx of Held-MNTB synapse. A hallmark of this synapse is the ability to maintain synaptic fidelity during high frequency transmission (i.e. at frequencies ? 800 Hz in vivo), a range at which conventional synaptic boutons cannot reliably fire. The nano-domain regulation of local ionic environments is a well-accepted signaling mechanism of synaptic transmission. Because of the high-fidelity necessary for the functional output of this synapse, extracellular synaptic K+ accumulation is likely higher at auditory synapses than traditional synaptic boutons during persistent spiking activity. We hypothesize that local microenvironments of K+ in the femtoliter volume of the synaptic space contributes to the activity-dependent regulation of STP at the presynaptic nerve terminal. The aims of this project are to 1) determine the homeostatic regulation of extracellular K+ during synaptic activity; 2) determine how postsynaptic depolarization regulates cytosolic Ca2+ concentration in the mature presynaptic nerve terminal; 3) determine how retrograde signaling modulates presynaptic STP. To pursue these aims, a combination of mouse genetics, synaptic electrophysiology and Ca2+ imaging approaches will be used. The results of this grant proposal will provide insights on the intrinsic modulation of synaptic strength and plasticity at developing and mature synapses of the auditory system. This will lead to a better understanding of healthy human spatial hearing and reveal potential synaptic mechanisms for treating hearing disorders.